Anionic Nucleotide−Lipids for In Vitro DNA Transfection

Abstract

A family of new anionic nucleotide based lipids featuring thymidine-3'-monophosphate as nucleotide and 1,2-diacyl-sn-glycerol as lipid moiety for in vitro delivery of nucleic acids is described. The nucleotide lipids were prepared in three steps starting from 1,2-diacyl-sn-glycerols and 2'-deoxythymidine-3'-phosphoramidite. Gel electrophoresis experiments show that nucleotide-based lipid-DNA complexes are observed at Ca(2+) concentration higher than 1 mM. The transfection experiments carried out on mammalian Hek cell lines clearly demonstrate that the nucleotide moiety enhances the transfection efficacy of the natural anionic DPPA and DPPG lipids. SAXS studies indicate that the enhancement in transfection for nucleotide-based lipid formulations compared to those of the abasic natural derivative (DPPA) is likely due to the presence of the 2D columnar inverted hexagonal phase (H(II)) with a unit cell parameter a = 69.1 A in the nucleotide lipid formulations. The cytotoxicity studies of lipoplexes, evaluated against Hek cells using an MTS assay, revealed that palmitoyl nucleotide derivative complexes were not toxic even after 4 h of incubation, thus indicating that the anionic nucleotide lipids presented in this work offer an alternative to cationic transfection reagents.

Advancing Nonviral Gene Delivery: Lipid- and Surfactant-Based Nanoparticle Design Strategies

“…This is accompanied by a shift in the zeta potential of these particles to positive values. Anionic complexes, having a negative zeta potential, have been successfully applied in a few cases [178,179]. The use of anionic complexes has been observed to reduce cytotoxicity of the resulting nanoparticles, possibly due to increased biocompatibility of the anionic lipids.…”

Designer lipids for drug delivery: From heads to tails

“…The hydrogen bonding and pi-stacking interactions of these nucleolipid overcame the repulsive force between the negative charge in the headgroup and the negative charge on nucleic acids in order to complex polyuridylic acid [96]. In a similar vein, Khiati and co-workers synthesized anionic nucleotide-lipids to decouple cationic interactions from H-bonding and pi-stacking (Figure 6F) [97]. Again, the forces of H-bonding and pi-stacking dominated the anionic repulsive forces between the nucleolipid and nucleic acid to complex and transfect eGFP into HEK cells in vitro [97].…”

“…In a similar vein, Khiati and co-workers synthesized anionic nucleotide-lipids to decouple cationic interactions from H-bonding and pi-stacking (Figure 6F) [97]. Again, the forces of H-bonding and pi-stacking dominated the anionic repulsive forces between the nucleolipid and nucleic acid to complex and transfect eGFP into HEK cells in vitro [97]. These studies show that electrostatic interactions are not necessary to complex and transfect nucleic acids in vitro , as H-bonding and pi-stacking forces can be sufficient.…”

An injectable liquid crystal system for sustained delivery of entecavir

“…To improve the sustained release pattern of the LCFS, DPPA was added to the LCFS. The DPPA is likely to be present as an anionic species at neutral pH conditions due to its pKa Khiati et al, 2009). On the contrary, entecavir is a slightly cationic drug that can form charge interactions with anionic species (Gao et al, 2014).…”